Peptide synthesis is the process by which amino acids are linked by amide bonds to produce peptides. The biological process of making long peptides, that is proteins, is known as protein photosynthesis.
Liquid-phase peptide synthesis is a classical approach to peptide synthesis and has been replaced in most labs by solid-phase synthesis. However, liquid-phase peptide synthesis retains usefulness in large-scale production of peptides for industrial purposes.
Solid-phase peptide synthesis (SPPS), is now the accepted method for creating peptides and proteins in the lab in a synthetic manner. SPPS allows the synthesis of natural peptides which are difficult to express in bacteria, the incorporation of unnatural amino acids, peptide/protein backbone modification, and the synthesis of D-proteins, which consist of D-amino acids. The process typically utilizes small solid insoluble porous beads which are treated with functional units on which peptide chains can be built. The resulting peptide chain will remains covalently attached to the bead until cleaved from that bead by a reagent such as anhydrous hydrogen fluoride or trifluoroacetic acid. The peptide is thus ‘immobilized’ on the solid-phase media or bead and can be retained during a filtration process, whereas liquid-phase reagents and by-products of synthesis are flushed away.
Repeated cycles of coupling-wash-deprotection-wash creates the desired peptide chain. The free N-terminal amine of a solid-phase attached peptide is coupled to a single N-protected amino acid unit. This unit is then deprotected, revealing a new N-terminal amine to which a further amino acid may be attached. The ability to perform wash cycles after each reaction provides a means to remove excess reagent with all peptide product remaining covalently attached to the insoluble resin bead. The objective is to generate high yield in each step. Thus each amino acid is added in major excess (2˜10×) and coupling amino acids together is optimized by the selection of agents. There are two major forms of SPPS utilized in labs and industry, Fmoc and Boc. Unlike ribosome protein synthesis, solid-phase peptide synthesis proceeds in a C-terminal to N-terminal fashion. The N-termini of amino acid monomers is protected by either of these two groups and added onto a deprotected amino acid chain.
SPPS is limited by yields, and typically peptides and proteins in the range of 70 amino acids are pushing the limits of synthetic accessibility. Synthetic difficulty also is sequence dependent and amyloid peptides and proteins are difficult to make.